Advanced ovarian cancer is a difficult condition for which new treatments are urgently needed. CD40 ligand (CD40L) is a candidate for the treatment of this disease because most ovarian cancer cells express the CD40 receptor. CD40L stimulation of this receptor kills the cancer cells, but has no cytotoxic effects on normal cells. In vivo, CD40L monotherapy markedly inhibited the growth of human ovarian cancer xenografts in immunodeficient mice. In combination with cisplatin, CD40L inhibited tumor growth by 99%. Of note, CD40L is also a key molecule in the immune system and activates dendritic cells to generate strong anti-tumor CD8+ T cell responses. Therefore, treating ovarian cancer with CD40L offers the possibility of a one-two punch: (1) Direct killing of CD40 receptor-positive ovarian cancer cells; and (2) Stimulation of anti-tumor CD8+ cytotoxic T cells. However, the crucial roadblock for moving CD40L into the clinic is that the production of 1st generation CD40L was dropped by its sole manufacturer. Multimeric Biotherapeutics now proposes to fill this void by producing a next-generation form of CD40L. The innovation is a highly active 2-trimer form of CD40L which is able to cluster the CD40 receptor, an essential requirement for CD40 cell signaling. This 2-trimer protein is made by genetically fusing the CD40L extracellular domain with a 2-armed scaffold protein (Acrp30) to make Acrp30-CD40L fusion protein (MegaCD40L). Three Aims are proposed: (1) To test murine MegaCD40L for its antitumor and immune activating effects in the ID8 mouse model of ovarian cancer; (2) To prepare human MegaCD40L that is active on human ovarian cancer cells in culture; and (3) To test the in vivo therapeutic effects of human MegaCD40L on human ovarian carcinoma xenografts in mice. These studies will provide the foundation for a follow-on Phase II SBIR project to prepare cGMP MegaCD40L suitable for clinical studies of ovarian and other cancers.